1. Field of the Invention
The invention relates generally to photodynamic therapy and more particularly, to the use of glyco-substituted dihydroxychlorins or glyco-substituted β-functionalized chlorin derivatives as photosensitizers for the treatment and prevention of microbial infectious diseases in human and animals.
2. State of the Art
Photodynamic therapy (PDT) is one of the most promising new techniques now being explored for use in a variety of medical applications, and particularly is a well-recognized treatment for the destruction of tumors. Photodynamic therapy uses light and a photosensitizer (a dye) to achieve its desired medical effect.
Antimicrobial photodynamic therapy is a very promising, relatively new method for combating bacterial infections even for resistant strains. Fortunately, no resistance to photodynamic destruction has been reported to be acquired by bacteria nor is it likely since the “killing species” is oxygen. Bacterial cells treated with photosensitizers were shown to be successfully killed by photo illumination. Due to the obvious differences between bacteria and malignant cells, photosensitizers with a different mode of action were needed for an antimicrobial PDT.
Effectiveness of the photosensitizers depends strongly on the bacterial cell wall as it becomes the limiting factor for the sensitizer penetration. While Gram-positive cells could be killed sufficiently by PDT, Gram-negative cells are more resistant to killing.
A major problem for the use of anti-microbial PDT is a blocking action of the components of the blood whose presence decreases the activity of photosensitizers. A high bactericidal photodynamic activity in PBS buffer could be decreased remarkably when blood serum or blood is added.
One of the prospective approaches to increase the specificity of photosensitizers and the effectiveness of PDT against bacterial infection is to conjugate a photosensitizer with a ligand-vector, which specifically binds to receptors on the surface of a target cell. In the prior art different methods have been used to effectively target the pathogen or infected cells.
U.S. Pat. No. 6,977,075 by Hasan et al. discloses a method of killing intracellular pathogens using antibiotics and PDT. The intracellular pathogens are targeted using conjugated photosensitizers. Targeting moiety used are molecules or a macromolecular structure that target macrophages or that interacts with a pathogen. Effectiveness of the conjugate against Gram-negative bacteria, and, in complex environment is not disclosed.
U.S. Pat. No. 6,573,258 by Bommer et al. describes cationic porphyrins which can effectively target both Gram-positive and Gram-negative bacteria when present at much lower concentrations and at much shorter irradiation times. The novel porphyrins have one hydrophobic tail consisting of at least one hydrocarbon chain of between 6 and 22 carbon in length. Bacterial targeting depends upon the carbon chain length and is not very effective.
U.S. Pat. No. 6,462,070 by Hasan et al. discloses a photosensitizer conjugated to polylysine which is linked to a histatin targeting moiety to treat disorder of the oral cavity infected by microorganism. These materials have trouble working in the presence of body fluids, such as saliva, blood, etc.
U.S. Pat. No. 5,466,681 describes a variety of conjugates useful for the treatment of infectious diseases due to pathogenic microorganisms. The conjugates comprise at least one agent coupled to a microorganism receptor—a carbohydrate vector, which is able to bind selectively to a microorganism. This patent discloses a conjugate comprising at least one agent that is an anti-infective, which couples to a microorganism receptor. Agents such as antibiotics, synthetic drugs and steroids are mentioned. Since photosensitizers do not themselves interact with microbes, they are not considered agents as described in this patent and were not disclosed therein.
Other promising approaches are about conjugates of porphyrins and carbohydrates. The publication: “Nitroglycosylated meso-arylporphyrins as Photoinhibitors of Gram positive Bacteria”, V. Sol, P. Branland, R. Granet, C. Kaldapa, B. Verneuil, P. Krausz, Bioorg. Med. Chem. Lett. 1998, 8, 3007-3010 describes the photodynamic activity of glycosylated nitroaryl-substituted porphyrins against Gram-positive bacteria, but no efficacy against Gram-negative bacteria and efficacy in complex environment is described. Moreover, the conjugates only show photodynamic activity when nitro-groups are present in the molecules.
Chlorins, as referred to in the present invention, are porphyrin derivatives, in which one double bond of the aromatic system in β-position is absent. Many current photosensitizers are not efficient enough as they have low absorption in the red region of the spectrum. Chlorins have the advantage that they possess an intense absorption in the red and near-infrared region of the electromagnetic spectrum which allows a deeper penetration of the light into the tissue. U.S. Pat. No. 7,022,843B1 by MacAlpine et discloses a variety of β,β′-dihydroxy meso-substituted chlorins as photosensitizers but do not offer guidance for treatment of microbial infectious diseases. Conversely, International Publication No. WO 2010/033678 by Wiehe et al. disclose unsymmetrically meso-substituted porphyrins and chlorins for diagnostic and PDT applications, including viral or infection diseases, however their effectiveness for both Gram-positive and Gram-negative bacteria in complex medium is not clearly conveyed.
There remains an urgent requirement to develop molecular conjugates which can actively target both Gram-positive and Gram-negative bacteria. Also they need to work under in vivo conditions, where typically blood and other body fluids are present, to use with patients directly to help protect them from deleterious microorganisms.